Fig 1.
pH profile of D. suzukii midgut.
The pH profile of the midgut of D. suzukii adults (7–8 days old) fed for 24 h on standard Drosophila diet containing either (a) bromocresol purple (0.5 w/w %) or (b) bromophenol blue (0.5 w/w %). In (a) purple indicates pH > 5.8 and yellow pH <5,2. In (b) yellow, pH < 3 and purple/blue, pH > 4.6. The central region of the midgut has a pH < 3, as indicated in the schematic (c).
Fig 2.
The size range of particles ingested by D. suzukii adults.
(a) A micrograph showing polydisperse suspension of PMMA particles dyed red with Sudan III ranging in size from 0.5–200 μm. (b) Dissected midgut of D. suzukii coloured red from ingested dyed particles but the opacity of gut hindered particle size determination. (c) Solid polymer particles in the fly excrement. The micrograph shows polymer particles resuspended from the excrement allowing particle size to be measured. (d) Comparison of the particle size range in the diet with that found in the excrement (n = 700). The maximum particle size excreted was ~ 15 μm, which we have used as an approximation for the ingestion limit. Particles < 5 μm were below the detection limit.
Fig 3.
Multiple emulsion template containing FITC-dextran (10 kDa).
Optical microscopy of the multiple emulsion microcapsule template prior to polymerisation (a and c) bright field images and (b and d) fluorescent images. The inner water droplets in the oil contain FITC-dextran that is localised in discrete pockets of fluorescence.
Fig 4.
Polymerised multiple emulsion template yielding solid P2VP microcapsules.
(a-d) Optical microscopy of microcapsules formed from polymerisation of the oil phase of the multiple emulsion template. (e) SEM analysis of the loaded microcapsules (f) Size measurement using light scattering, average distribution expressed as volume (%) (n = 10), of the microcapsules ranging from 2–100 μm. The smaller peak around 1 μm is a by-product formed of solid polymer particles containing no active species.
Fig 5.
pH dependence of P2VP microcapsule dissolution.
(a) Determination of the pKa of the microcapsules undertaken by monitoring light transmission (500 nm) as a function of pH. Data (mean±s.e.m; n = 3) were fitted to a sigmoidal curve using OriginPro 9.1 software. (b) Images taken at 0, 3, 6 and 12 sec from a video recording showing the effect of reducing pH on the P2VP microcapsules (see methods section) by introducing citric acid. The reduction in pH leads to swelling and rapid dissolution of the microcapsules, releasing their cargo.
Fig 6.
In vitro release of FITC-dextran from responsive microcapsules in different pH buffers.
P2VP pH-responsive microcapsules in a pH 6 (n = 4) environment showed no release of encapsulated FITC-dextran over 24 h as the pH was above the pKa of the microcapsules. At the lower pH of 3.8 (n = 4) around 80% of the encapsulated FITC-dextran was released within the first 5 mins, no further release was observed after this time. It is assumed that the unaccounted 20% is lost during the production of the microcapsules.
Fig 7.
In vivo release of FITC-dextran triggered by pH changes in D. suzukii gut.
In vivo release of FITC-dextran from not from control non–responsive PMMA (a-c), but pH-responsive P2VP (d-f) microcapsules fed to D. suzukii adults. Intact fluorescent PMMA microcapsules in the crop (a), excrement (b) and the lumen of the midgut (c). Intact pH-responsive P2VP microcapsules in the crop (d), but none in the excrement (e) and midgut (f). Dispersed fluorescence in the posterior midgut shows that the P2VP microcapsules have released their contents. Fluorescence was also seen in the haemolymph released from the body during dissection. Strong fluorescence accumulated in the Malpighian tubules (Mt) demonstrating uptake of FITC-dextran from the haemolymph. All scale bars 200 μm. (g) Fluorescence in the extracted haemolymph of flies fed normal diet (control), normal diet with PMMA non-responsive microcapsules and normal diet with P2VP pH-responsive microcapsules (mean±s.e.m; n = 3). The fluorescence is significantly more intense in the haemolymph of flies fed responsive P2VP microcapsules compared to the control and non-responsive PMMA microcapsules (P = 0.0004).